An Electrospray Ionization−Flow Tube Study of H/D Exchange in Protonated Bradykinin

An electrospray ionization−fast flow technique has been employed to study the reactions of doubly protonated bradykinin and des-Arg 9 -bradykinin with CH 3 OD and ND 3, respectively. Deconvolution of the experimental mass spectral data followed by simulation of the kinetic data by solution of differential equations leads to sets of apparent and site-specific rate constants. On a time scale of several milliseconds, bradykinin undergoes with ND 3 three fast H/D exchanges and one slow exchange. Three equivalent exchanges are observed with CH 3 OD that are nearly 2 orders of magnitude slower than the ND 3 reactions. Up to six hydrogen exchanges are observed for the reaction of des-Arg 9 -bradykinin with ND 3 . The more efficient exchange of des-Arg 9 -bradykinin is accompanied by formation of collisionally stabilized complexes between doubly protonated des-Arg 9 -bradykinin and ND 3 at a He carrier gas pressure of about 0.2 Torr. Multiple-collision activation−collision-induced dissociation of reactant and product ions of the isotope exchange reactions was carried out in front of the sampling nose cone of the analyzer quadrupole mass filter system. The degree of deuterium incorporation into the parent doubly protonated ions and into several of the b n + and y n + ions combined with the site-specific rate constants obtained indicates that the three equivalent hydrogens exchanged in doubly protonated bradykinin are at the protonated N-terminus amine group. Complexation of doubly protonated bradykinin by ND 3 is prevented by its tightly folded structure, and this in turn prevents H/D exchange of the amide hydrogens of bradykinin. The additional H/D exchanges observed in the case of doubly protonated des-Arg 9 -bradykinin are made possible by complexation of its less compact structure via hydrogen-bonded intermediates that promote H/D exchange of amide hydrogens.